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Deep Space Nine went where no Star Trek had gone before – it was the first series that was not based on a starship, but was instead based on a starbase, known as Deep Space 9 (the starship USS Defiant was introduced in season 3, but the station remained the primary setting of the series). Indeed, when Brandon Tartikoff originally approached Rick Berman about the show, he specifically said he wanted it to have a new format; if The Next Generation was Wagon Train in space, Deep Space Nine was to be The Rifleman in space – a man and his son coming to a dilapidated town on the edge of a new frontier.

The series was designed to have more interpersonal conflict than its predecessors, while still staying true to the universe that Gene Roddenberry had created. Rick Berman commented: "[Deep Space 9]'s an alien space station that doesn't work the way they want it to, and that in itself created a lot of conflict. At the same, our core characters are Starfleet officers, Sisko, O'Brien, the doctor and Dax in no way vary from The Next Generation in terms of the lack of conflict among themselves. That was a rule we had to follow. We needed to create a series that wasn't a franchise based on people aboard a starship, because we knew there would a couple of years of overlap between the two series". (Captains' Logs Supplemental - The Unauthorized Guide to the New Trek Voyages, p 5)

Berman also commented: "The problem with Star Trek: The Next Generation is Gene created a group of characters that he purposely chose not to allow conflict between. Starfleet officers cannot be in conflict, thus its murderous to write these shows because there is no good drama without conflict, and the conflict has to come from outside the group. What we wanted to do was something that was almost paradoxical - bring conflict but not break Gene's rules. They still play paramount importance in what we're doing. We created an environment where Starfleet officers were in a location that they weren't happy about being in, and they were in a location where the people who lived there weren't all that happy about them being there. We also created a situation where we had people who were members of our core group who were not Starfleet: the security shapeshifter Odo; the Bajoran Major, Kira; the bartender, Quark. A group of our integral people are not Starfleet officers, and the ones that are Starfleet officers aren't crazy about where they are, so we have a lot of frustration and conflict". (Captains' Logs Supplemental - The Unauthorized Guide to the New Trek Voyages, p 8)

Regarding Gene Roddenberry's involvement, Berman stated, "Michael (Piller) and I discussed it with Gene when we were still in the early stages, but never anything conceptual.""We never got a chance to discuss it (the concept) with Gene. By the time we had it to the point that it was discussable, he was in pretty bad shape and not really in the condition that it would have been wise to discuss it with him. On two specific occasions I was with him at his house and we tried to bring it up, but it wasn't really appropriate." (Captains' Logs: The Unauthorized Complete Trek Voyages, p. 328)

In addition to the visits to the mirror universe, the DS9 writing staff wrote a number of episodes where the character of Miles O'Brien would be subject to particular trauma. This became an in-joke among the staff, who called them "O'Brien Must Suffer" episodes and went to great lengths to produce at least one such episode per season. (Star Trek: Deep Space Nine Companion)

The show also focused on a wider array of uses and depictions of functions for the holodeck. In addition to many obvious activities such as those referenced by Chief O'Brien and Julian Bashir, which were completely in keeping with holodeck usage on ST:TNG, the holodeck was used as a recurring background for people to hang out in in the form of a 1960s Las Vegas lounge (numerous episodes), as a weapons showroom (by Quark), as well as the location for a baseball game between teams assembled by Sisko vs. Captain Solok, a long-time rival Vulcan captain ("Take Me Out to the Holosuite").

The show broke the "standard format" for Star Trek shows a number of times as well, with a direct, first-person narrative providing the commentary for the episode "In the Pale Moonlight", a retelling of a classic TOS episode from a different angle in "Trials and Tribble-ations", life in the racially segregated 1950s in "Far Beyond the Stars", and brought the concept of "Black Ops" to the Star Trek Universe with Section 31: "Inquisition". The show also broke with tradition - and with the two Star Trek series that followed it - by featuring a commanding officer as the star of the show at the rank of commander, rather than captain, for a significant portion of its run. Robert Hewitt Wolfe recalled that this led to unfavorable comparisons to the other series. "Whenever people would do articles about Star Trek they would talk about the three captains: Kirk, Picard, and Janeway." Feeling that Sisko deserved the higher rank as much as the other lead characters, the producers decided to promote Sisko in "The Adversary". (Star Trek: Deep Space Nine Companion, p. 253)

Due to the show's non-episodic nature, much of the series was easily lost on the casual viewer. Many also believe that the changing television landscape contributed to DS9's ratings trouble, as local TV stations which had aired TNG in prime time became WB and UPN affiliates and pushed syndicated programming to the margins. Subsequent Trek shows Star Trek: Voyager and Star Trek: Enterprise had network support from UPN and a guaranteed time slot. DS9 was also the only series to run opposite another StarTrek show (first The Next Generation, then Voyager) for the entirety of its run (the first 12 episodes of the third season aired without another series on).

Additionally, certain markets, notably in the UK, would only play one Star Trek series, in its entirety, at a time. Thus, events alluded to in The Next Generation or Voyager that happened in Deep Space Nine took months to "sync up."

Despite these problems, Deep Space Nine remained a fan-favorite series throughout its seven-year run, with reviewers consistently lauding the series for its bold shift in tone from The Next Generation. Most notable among such changes was the concept of inter-personal conflict – something which Gene Roddenberry himself was said to have forbidden.

"I'd like us to be remembered as the Trek series that dared to be different.
We took chances in a franchise that has every reason to play it safe and
spoon-feed the same old thing to the audience week after week. We
challenged the characters, the audience, and the Star Trek universe itself.
Sometimes we failed (sometimes spectacularly) but we never stopped trying to
push the show into new directions. - Ronald D. Moore DS9 producer, screenwriter"

There was also a rivalry with another popular and critically acclaimed television series, Babylon 5, created and produced by J. Michael Straczynski for Warner Bros. The two productions, which ran largely concurrently, were observed to be so similar that Babylon 5 fans accused Paramount, to whom Straczynski had previously pitched his series, of plagiarism. Considering how fellow Trek alumni like Walter Koenig and Andreas Katsulas had major roles in the rival series, Majel Barrett-Roddenberry agreed to a guest appearance as a gesture of goodwill to encourage a reconciliation between the fandoms.

Deep Space Nine was the first live-action Star Trek series to feature a fully-animated sequence in its opening credits, as opposed to the simple flashcards accompanied by rapid flypasts of the Enterprise used for the opening sequences of both The Original Series and The Next Generation. Both subsequent series, Voyager and Enterprise would also have fully animated credit sequences.

In the DVD season special features the creators talk about the similarities between Deep Space Nine and an old Western setting Sisko as the Mayor, Kira as a Native American, Bashir as a country doctor, Odo the lawman, Quark as the local barkeep and Miles O'Brien the everyman wed to the local school marm Keiko.